Project Summary/Abstract Over 1,000,000 endovascular procedures are performed in the United States each year and restenosis continues to be the leading cause of failure. In animal models, neointimal development after balloon injury resembles restenosis in humans. Despite major research advances using this and other models, our ability to translate our knowledge into clinically relevant therapies has not been realized. One factor that has increasingly been associated with accelerated restenosis after endovascular interventions is the presence of calcification in the arterial wall. During arterial calcification, smooth muscle cells (SMCs) in the media undergo phenotypic transformation into a more osteochondrogenic cell type. The effects of calcifying SMCs on neointimal formation have not been assessed. The bone morphogenetic proteins (BMPs) are a family of signaling molecules that were named for their ability to induce ectopic bone formation. We have previously shown that the BMP4 inhibits SMC proliferation. Medial cells in calcified arteries, however, are more similar to those in developing bone where cells display an exuberant proliferative response to BMP4 in both in vitro and in vivo studies. Based on these data, we hypothesize that accelerated restenosis in calcified arteries occurs due to enhanced migration, proliferation, and matrix synthesis of calcifying SMCs through a mechanism involving BMP4-BMP-receptor signaling. In order to test this hypothesis, we will perform the following specific aims: 1) Compare the effects of BMP4 on migration, proliferation, and matrix synthesis in calcified versus uncalcified SMCs; 2) Determine whether BMP-receptor-I activation accelerates injury-induced neointimal development in calcified arteries compared with uncalcified arteries; 3) Determine whether blocking BMP-receptor-I activation using a small molecule inhibitor can reduce neointimal formation after injury in calcified arteries. In order to address these aims, we will take an innovative approach - manipulation of the SMC phenotype prior to injury - to evaluate the effects of arterial calcification on neointimal development after balloon injury. In completing these aims we will answer a fundamental question about the behavior of SMCs in calcified arteries. Namely, we will determine if their osteochondrogenic transformation causes them to respond to injury more like cells in developing bone than normal SMCs. If our hypothesis is correct, it will suggest that therapies to prevent restenosis must be tailored to the particular phenotype of cells in the arterial wall. Additionally, during these experiments we will evaluate a novel, small-molecule BMP inhibitor which may have significant clinical relevance for preventing restenosis in patients with peripheral artery disease.